Characterization of a new 2.4-kb plasmid of Corynebacterium casei and development of stable corynebacterial cloning vector.
A new plasmid pCASE1 was isolated from Gram-positive Corynebacterium casei JCM 12072. It comprised a 2.4-kb nucleotide sequence with three ORFs, two of which were indispensable for autonomous replication in Corynebacterium glutamicum. Homology search identified these two ORFs as repA and repB, areas coding proteins involved in plasmid replication. repA sequence showed high similarity to theta-replicating Escherichia coli ColE2-P9 plasmids and even higher similarity to plasmids derived from Gram-positive bacteria belonging to a subfamily of this ColE2-P9 group. An E. coli-C. glutamicum shuttle vector was constructed with pCASE1 fragment including repA and repB to transform C. glutamicum and showed compatibility with corynebacterial plasmids from different plasmid families. The copy number of the shuttle vector in C. glutamicum was 13 and the vector showed stability for 102 generations with no selective pressure.
Tsuchida
Yoshiki
Y
Research Institute of Innovative Technology for the Earth, 9-2 Kizugawadai, Kizugawa, Kyoto 619-0292, Japan.
Kimura
Sakurako
S
Suzuki
Nobuaki
N
Inui
Masayuki
M
Yukawa
Hideaki
H
D003352:Corynebacterium / Q000235:genetics*; D048230:Corynebacterium glutamicum / Q000235:genetics; D004261:DNA Replication; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D005822:Genetic Vectors; D005827:Genetics, Microbial / Q000379:methods*; D008967:Molecular Biology / Q000379:methods*; D016366:Open Reading Frames; D010957:Plasmids; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D004269:DNA, Bacterial